Innate immune cells, such as macrophages, express germ-line encoded pattern recognition receptors that recognize conserved pathogen-associated patterns (PAMPs) on microbes. The Toll-like receptor family (TLR) is the prototypic family of pattern recognition receptors. Humans express ten known TLRs and the ligands for a subset have been identified. For example, TLR4 recognizes bacterial lipopolysaccharides, and TLR2 combined with TLR6 recognizes peptidoglycan. TLRs trigger both shared and receptor-specific inflammatory signals. Thus both TLR4 and TLR2/6 trigger shared signals such as those leading to NF-kB activation and the production of TNF-alpha, whereas TLR4 also activates specific signals such as the induction of IP-10. Therefore, specific recognition of PAMPs by TLRs results in a "tailored" inflammatory response that is appropriate to the infection. Many pathogenic bacteria are flagellated, and we recently identified flagellin, the major component of flagella, as the stimulus that activates TLR5. The focus of this proposal is to determine the precise mechanism by which TLR5 recognizes flagellin, and how this initiates an appropriate inflammatory response. We aim to 1) identify the site on flagellin that is recognized by TLR5, 2) characterize the physical interaction between TLR5 and flagellin, 3) determine the cellular and tissue distribution of TLR5, 4) analyze TLR5-dependent regulation of gene expression, and 5) characterize functional domains of the TLR5 cytoplasmic tail that confer either general or specific inflammatory signals. This study will substantially expand our understanding of innate immunity. The emerging model is that different TLRs decipher a "bar code" on the invading pathogen and then trigger a composite set of signals that result in an inflammatory response that precisely orchestrates host defense to the specific organism.